Lifespan Modulation in Mice and the Confounding Effects of Genetic Background

We are currently in the midst of a revolution in ageing research,with several dietary,genetic and pharmacological interventions now known to modulate ageing in model organisms.Excitingly,these interventions also appear to have beneficial effects on late-life health.For example,dietary restriction(DR) has been shown to slow the incidence of age-associated cardiovascular disease,metabolic disease,cancer and brain ageing in non-human primates and has been shown to improve a range of health indices in humans.While the idea that DR's ability to extend lifespan is often thought of as being universal,studies in a range of organisms,including yeast,mice and monkeys,suggest that this may not actually be the case.The precise reasons underlying these differential effects of DR on lifespan are currently unclear,but genetic background may be an important factor in how an individual responds to DR.Similarly,recent findings also suggest that the responsiveness of mice to specific genetic or pharmacological interventions that modulate ageing may again be influenced by genetic background.Consequently,while there is a clear driver to develop interventions to improve late-life health and vitality,understanding precisely how these act in response to particular genotypes is critical if we are to translate these findings to humans.We will consider of the role of genetic background in the efficacy of various lifespan interventions and discuss potential routes of utilising genetic heterogeneity to further understand how particular interventions modulate lifespan and healthspan.     

The Relationship between Intelligence and Training Gains Is Moderated by Training Strategy

We examined the relationship between training regimen and fluid intelligence in the learning of a complex video game. Fifty non-game-playing young adults were trained on a game called Space Fortress for 30 hours with one of two training regimens: 1) Hybrid Variable-Priority Training (HVT), with part-task training and a focus on improving specific skills and managing task priorities, and 2) Full Emphasis Training (FET) in which participants practiced the whole game to obtain the highest overall score. Fluid intelligence was measured with the Raven’s Progressive Matrix task before training. With FET, fluid intelligence was positively associated with learning, suggesting that intellectual ability played a substantial role in determining individual differences in training success. In contrast, with HVT, fluid intelligence was not associated with learning, suggesting that individual differences in fluid intelligence do not factor into training success in a regimen that emphasizes component tasks and flexible task coordination. By analyzing training effects in terms of individual differences and training regimens, the current study offers a training approach that minimizes the potentially limiting effect of individual differences.     

The Requirement of WHIRLY1 for Embryogenesis Is Dependent on Genetic Background in Maize

Plastid gene expression is essential to embryogenesis in higher plants, but the underlying mechanism is obscure. Through molecular characterization of an embryo defective 16 (emb16) locus, here we report that the requirement of plastid translation for embryogenesis is dependent on the genetic background in maize (Zea mays). The emb16 mutation arrests embryogenesis at transition stage and allows the endosperm to develop largely normally. Molecular cloning reveals that Emb16 encodes WHIRLY1 (WHY1), a DNA/RNA binding protein that is required for genome stability and ribosome formation in plastids. Interestingly, the previous why1 mutant alleles (why1-1 and why1-2) do not affect embryogenesis, only conditions albino seedlings. The emb16 allele of why1 mutation is in the W22 genetic background. Crosses between emb16 and why1-1 heterozygotes resulted in both defective embryos and albino seedlings in the F1 progeny. Introgression of the emb16 allele from W22 into A188, B73, Mo17, Oh51a and the why1-1 genetic backgrounds yielded both defective embryos and albino seedlings. Similar results were obtained with two other emb mutants (emb12 and emb14) that are impaired in plastid protein translation process. These results indicate that the requirement of plastid translation for embryogenesis is dependent on genetic backgrounds, implying a mechanism of embryo lethality suppression in maize.     

Genic-SSR对中国兰的遗传背景分析

本研究利用本实验室开发的62个genic-SSR标记分析5个中国兰种的43个品种,其中52个标记呈多态性,其多态性指数(PIC)从0.054到0.824不等,平均为0.415,遗传多样性分析显示这43个材料之间的遗传距离平均为0.358,范围为0.016~0.600。对这些材料进行聚类分析和主成分分析,结果表明大多数来自一个物种的中国兰品种聚类在一起,而春兰分别聚类于几个系统树分支上,具有多起源性。与地理分布信息交叉分析发现:同一物种的品种往往聚类于一个大的系统树分支,但来自于不同地区的品种分别聚类于不同小分支;一些来自临近地域的品种往往聚类在一起,而这些品种有时不属于同一物种。这些结果说明遗传背景和地理环境存在一定的联系。     

显微注射法制备遗传工程小鼠模型的研究

遗传工程小鼠模型是基因功能、人类疾病发病机制及新药研究开发的最重要的模式生物之一。在建立遗传工程小鼠模型的众多方法中,显微注射法是目前国际上公认的制备转基因及基因剔除动物模型的首选。在进行了大量显微注射工作后,简要介绍建立遗传工程小鼠模型的基本技术与方法,并对在显微操作过程中较为关键的因素进行一些分析和讨论。     

Rapid Decline of Ceftazidime Resistance in Antibiotic-Free and Sublethal Environments Is Contingent on Genetic Background

Trade-offs of antibiotic resistance evolution, such as fitness cost and collateral sensitivity (CS), could be exploited to drive evolution toward antibiotic susceptibility. Decline of resistance may occur when resistance to other drug leads to CS to the first one and when compensatory mutations, or genetic reversion of the original ones, reduce fitness cost. Here we describe the impact of antibiotic-free and sublethal environments on declining ceftazidime resistance in different Pseudomonas aeruginosa resistant mutants. We determined that decline of ceftazidime resistance occurs within 450 generations, which is caused by newly acquired mutations and not by reversion of the original ones, and that the original CS of these mutants is preserved. In addition, we observed that the frequency and degree of this decline is contingent on genetic background. Our results are relevant to implement evolution-based therapeutic approaches, as well as to redefine global policies of antibiotic use, such as drug cycling.     

Temporal and Anatomical Host Resistance to Chronic Salmonella Infection Is Quantitatively Dictated by Nramp1 and Influenced by Host Genetic Background

The lysosomal membrane transporter, Nramp1, plays a key role in innate immunity and resistance to infection with intracellular pathogens such as non-typhoidal Salmonella (NTS). NTS-susceptible C57BL/6 (B6) mice, which express the mutant Nramp1^D169 allele, are unable to control acute infection with Salmonella enterica serovar Typhimurium following intraperitoneal or oral inoculation. Introducing functional Nramp1^G169 into the B6 host background, either by constructing a congenic strain carrying Nramp1^G169 from resistant A/J mice (Nramp-Cg) or overexpressing Nramp1^G169 from a transgene (Nramp-Tg), conferred equivalent protection against acute Salmonella infection. In contrast, the contributions of Nramp1 for controlling chronic infection are more complex, involving temporal and anatomical differences in Nramp1-dependent host responses. Nramp-Cg, Nramp-Tg and NTS-resistant 129×1/SvJ mice survived oral Salmonella infection equally well for the first 2–3 weeks, providing evidence that Nramp1 contributes to the initial control of NTS bacteremia preceding establishment of chronic Salmonella infection. By day 30, increased host Nramp1 expression (Tg>Cg) provided greater protection as indicated by decreased splenic bacterial colonization (Tg<Cg). However, despite controlling bacterial growth within MLN as effectively as 129×1/SvJ mice, Nramp-Cg and Nramp-Tg mice eventually succumbed to infection. These data indicate: 1) discrete, anatomically localized host resistance is conferred by Nramp1 expression in NTS-susceptible mice, 2) restriction of systemic bacterial growth in the spleens of NTS-susceptible mice is enhanced by Nramp1 expression and dose-dependent, and 3) host genes other than Nramp1 also contribute to the ability of NTS-resistant 129×1/SvJ mice to control bacterial replication during chronic infection.     

The Influence of Genetic Background and the Homologous Chromosome 17 on t-Haplotype Transmission Ratio Distortion in Mice

Transmission ratio distortion is a characteristic of complete t-haplotypes, such that heterozygous males preferentially transmit the t-haplotype bearing chromosome 17 to the majority of their progeny. At least two genes contained within the t-haplotype have been identified as being required for such high transmission ratios. In this study we examine the effects of the genetic background and the chromosome homologous to the t-haplotype on transmission ratio distortion. We use two different congenic lines: BTBRTF/Nev.Ttf/t12, in which the t12 haplotype has a transmission ratio of 52%, and C3H/DiSn.Ttf/t12, in which the t12 haplotype has a transmission ratio of 99%. By intercrossing these two strains to produce reciprocal F1 and F2 generations, we have isolated the effects of the homologous chromosome 17 from the effects of the genetic background. We demonstrate that both the homologous chromosome and the genetic background have profound effects on t-haplotype transmission ratio distortion. Furthermore, it is evident that the t-haplotype transmission ratio behaves as a quantitative character rather than an intrinsic property of t-haplotypes.     

Effects of Vendor and Genetic Background on the Composition of the Fecal Microbiota of Inbred Mice

The commensal gut microbiota has been implicated as a determinant in several human diseases and conditions. There is mounting evidence that the gut microbiota of laboratory mice (Mus musculus) similarly modulates the phenotype of mouse models used to study human disease and development. While differing model phenotypes have been reported using mice purchased from different vendors, the composition and uniformity of the fecal microbiota in mice of various genetic backgrounds from different vendors is unclear. Using culture-independent methods and robust statistical analysis, we demonstrate significant differences in the richness and diversity of fecal microbial populations in mice purchased from two large commercial vendors. Moreover, the abundance of many operational taxonomic units, often identified to the species level, as well as several higher taxa, differed in vendor- and strain-dependent manners. Such differences were evident in the fecal microbiota of weanling mice and persisted throughout the study, to twenty-four weeks of age. These data provide the first in-depth analysis of the developmental trajectory of the fecal microbiota in mice from different vendors, and a starting point from which researchers may be able to refine animal models affected by differences in the gut microbiota and thus possibly reduce the number of animals required to perform studies with sufficient statistical power.     

Graft-versus-Host Disease Is Enhanced by Selective CD73 Blockade in Mice

CD73 functions as an ecto-5′-nucleotidase to produce extracellular adenosine that has anti-inflammatory and immunosuppressive activity. We here demonstrate that CD73 helps control graft-versus-host disease (GVHD) in mouse models. Survival of wild-type (WT) recipients of either allogeneic donor naïve CD73 knock-out (KO) or WT T cells was similar suggesting that donor naïve T cell CD73 did not contribute to GVHD. By contrast, donor CD73 KO CD4⁺CD25⁺ regulatory T cells (Treg) had significantly impaired ability to mitigate GVHD mortality compared to WT Treg, suggesting that CD73 on Treg is critical for GVHD protection. However, compared to donor CD73, recipient CD73 is more effective in limiting GVHD. Pharmacological blockade of A2A receptor exacerbated GVHD in WT recipients, but not in CD73 KO recipients, suggesting that A2 receptor signaling is primarily implicated in CD73-mediated GVHD protection. Moreover, pharmacological blockade of CD73 enzymatic activity induced stronger alloreactive T cell activity, worsened GVHD and enhanced the graft-versus-leukemia (GVL) effect. These findings suggest that both donor and recipient CD73 protects against GVHD but also limits GVL effects. Thus, either enhancing or blocking CD73 activity has great potential clinical application in allogeneic bone marrow transplants.     

BDNF Val66Met Polymorphism Is Associated with Self-Reported Empathy

Empathy is an important driver of human social behaviors and presents genetic roots that have been studied in neuroimaging using the intermediate phenotype approach. Notably, the Val66Met polymorphism of the Brain-derived neurotrophic factor (BDNF) gene has been identified as a potential target in neuroimaging studies based on its influence on emotion perception and social cognition, but its impact on self-reported empathy has never been documented. Using a neurogenetic approach, we investigated the association between the BDNF Val66Met polymorphism and self-reported empathy (Davis’ Interpersonal Reactivity Index; IRI) in a sample of 110 young adults. Our results indicate that the BDNF genotype is significantly associated with the linear combination of the four facets of the IRI, one of the most widely used self-reported empathy questionnaire. Crucially, the effect of BDNF Val66Met goes beyond the variance explained by two polymorphisms of the oxytocin transporter gene previously associated with empathy and its neural underpinnings (OXTR rs53576 and rs2254298). These results represent the first evidence suggesting a link between the BDNF gene and self-reported empathy and warrant further studies of this polymorphism due to its potential clinical significance.     

Genetic Signature of Histiocytic Sarcoma Revealed by a Sleeping Beauty Transposon Genetic Screen in Mice

Histiocytic sarcoma is a rare, aggressive neoplasm that responds poorly to therapy. Histiocytic sarcoma is thought to arise from macrophage precursor cells via genetic changes that are largely undefined. To improve our understanding of the etiology of histiocytic sarcoma we conducted a forward genetic screen in mice using the Sleeping Beauty transposon as a mutagen to identify genetic drivers of histiocytic sarcoma. Sleeping Beauty mutagenesis was targeted to myeloid lineage cells using the Lysozyme2 promoter. Mice with activated Sleeping Beauty mutagenesis had significantly shortened lifespan and the majority of these mice developed tumors resembling human histiocytic sarcoma. Analysis of transposon insertions identified 27 common insertion sites containing 28 candidate cancer genes. Several of these genes are known drivers of hematological neoplasms, like Raf1, Fli1, and Mitf, while others are well-known cancer genes, including Nf1, Myc, Jak2, and Pten. Importantly, several new potential drivers of histiocytic sarcoma were identified and could serve as targets for therapy for histiocytic sarcoma patients.     

eNOS Activation by HDL Is Impaired in Genetic CETP Deficiency

Mutations in the CETP gene resulting in defective CETP activity have been shown to cause remarkable elevations of plasma HDL-C levels, with the accumulation in plasma of large, buoyant HDL particles enriched in apolipoprotein E. Genetic CETP deficiency thus represents a unique tool to evaluate how structural alterations of HDL impact on HDL atheroprotective functions. Aim of the present study was to assess the ability of HDL obtained from CETP-deficient subjects to protect endothelial cells from the development of endothelial dysfunction. HDL isolated from one homozygous and seven heterozygous carriers of CETP null mutations were evaluated for their ability to down-regulate cytokine-induced cell adhesion molecule expression and to promote NO production in cultured endothelial cells. When compared at the same protein concentration, HDL and HDL3 from carriers proved to be as effective as control HDL and HDL3 in down-regulating cytokine-induced VCAM-1, while carrier HDL2 were more effective than control HDL2 in inhibiting VCAM-1 expression. On the other hand, HDL and HDL fractions from carriers of CETP deficiency were significantly less effective than control HDL and HDL fractions in stimulating NO production, due to a reduced eNOS activating capacity, likely because of a reduced S1P content. In conclusion, the present findings support the notion that genetic CETP deficiency, by affecting HDL particle structure, impacts on HDL vasculoprotective functions. Understanding of these effects might be important for predicting the outcomes of pharmacological CETP inhibition.     

多重PCR在几个近交系小鼠遗传检测中的应用初探

目的　探讨多重PCR方法在小鼠微卫星检测中的应用。方法　用 34对特异性引物对AKR、BALB c、C57 BL、DBA 2、CBA、A WY、6 15、T739、BALB cJ和AKR J 10个品系的近交系小鼠用PCR方法进行遗传检测 ,并从中选用 4对引物 ,对这些品系小鼠进行二重和多重PCR检测。结果　二重PCR在与单一PCR相同的反应条件下 ,扩增出两条与预期条带相同的带 ,而三重PCR则没有得到三条预期的条带 ,出现了干扰现象。结论　通过二条条带的距离可以鉴别出不同的近交系小鼠 ,二重PCR可应用于近交系小鼠的遗传检测 ,具有方便简单、省时的优点     

Low Bone Strength Is a Manifestation of Phenylketonuria in Mice and Is Attenuated by a Glycomacropeptide Diet

Purpose

Phenylketonuria (PKU), caused by phenylalanine (phe) hydroxylase loss of function mutations, requires a low-phe diet plus amino acid (AA) formula to prevent cognitive impairment. Glycomacropeptide (GMP), a low-phe whey protein, provides a palatable alternative to AA formula. Skeletal fragility is a poorly understood chronic complication of PKU. We sought to characterize the impact of the PKU genotype and dietary protein source on bone biomechanics.

Procedures

Wild type (WT; Pah^+/+) and PKU (Pah^enu2/enu2) mice on a C57BL/6J background were fed high-phe casein, low-phe AA, and low-phe GMP diets between 3 to 23 weeks of age. Following euthanasia, femur biomechanics were assessed by 3-point bending and femoral diaphyseal structure was determined. Femoral ex vivo bone mineral density (BMD) was assessed by dual-enengy x-ray absorptiometry. Whole bone parameters were used in prinicipal component analysis. Data were analyzed by 3-way ANCOVA with genotype, sex, and diet as the main factors.

Findings

Regardless of diet and sex, PKU femora were more brittle, as manifested by lower post-yield displacement, weaker, as manifested by lower energy and yield and maximal loads, and showed reduced BMD compared with WT femora. Four principal components accounted for 87% of the variance and all differed significantly by genotype. Regardless of genotype and sex, the AA diet reduced femoral cross-sectional area and consequent maximal load compared with the GMP diet.

Conclusions

Skeletal fragility, as reflected in brittle and weak femora, is an inherent feature of PKU. This PKU bone phenotype is attenuated by a GMP diet compared with an AA diet.     

A Specific Nuclear DNA Background Is Required for High Frequency Lymphoma Development in Transmitochondrial Mice with G13997A mtDNA

We previously found that mouse mitochondrial DNA (mtDNA) with a G13997A mutation (G13997A mtDNA) controls not only the transformation of cultured lung carcinoma cells from poorly metastatic into highly metastatic cells, but also the transformation of lymphocytes into lymphomas in living C57BL/6 (B6) mice. Because the nuclear genetic background of the B6 strain makes the strain prone to develop lymphomas, here we examined whether G13997A mtDNA independently induces lymphoma development even in mice with the nuclear genetic background of the A/J strain, which is not prone to develop lymphomas. Our results showed that the B6 nuclear genetic background is required for frequent lymphoma development in mice with G13997A mtDNA. Moreover, G13997A mtDNA in mice did not enhance the malignant transformation of lung adenomas into adenocarcinomas or that of hepatocellular carcinomas from poorly metastatic into highly metastatic carcinomas. Therefore, G13997A mtDNA enhances the frequency of lymphoma development under the abnormalities in the B6 nuclear genome, and does not independently control tumor development and tumor progression.     

Genetic Diversity in RNA Virus Quasispecies Is Controlled by Host-Virus Interactions

Many RNA viruses have genetically diverse populations known as quasispecies. Important biological characteristics may be related to the levels of diversity in the quasispecies (quasispecies cloud size), including adaptability and host range. Previous work using Tobacco mosaic virus and Cucumber mosaic virus indicated that evolutionarily related viruses have very different levels of diversity in a common host. The quasispecies cloud size for these viruses remained constant throughout serial passages. Inoculation of these viruses on a number of hosts demonstrated that quasispecies cloud size is not constant for these viruses but appears to be dependent on the host. The quasispecies cloud size remained constant as long as the viruses were maintained on a given host. Shifting the virus between hosts resulted in a change in cloud size to levels associated with the new host. Quasispecies cloud size for these viruses is related to host-virus interactions, and understanding these interactions may facilitate the prediction and prevention of emerging viral diseases.